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A millifluidic study of cell-to-cell heterogeneity in growth-rate and cell-division capability in populations of isogenic cells of Chlamydomonas reinhardtii.

Damodaran SP, Eberhard S, Boitard L, Rodriguez JG, Wang Y, Bremond N, Baudry J, Bibette J, Wollman FA - PLoS ONE (2015)

Bottom Line: We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers) and a significant subpopulation of slowly dividing cells (slow-growers).Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times.We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Colloïdes et Matériaux Divisés, Institute of Chemistry, Biology and Innovation ESPCI ParisTech/CNRS UMR 8231/PSL* Research University, Paris, France.

ABSTRACT
To address possible cell-to-cell heterogeneity in growth dynamics of isogenic cell populations of Chlamydomonas reinhardtii, we developed a millifluidic drop-based device that not only allows the analysis of populations grown from single cells over periods of a week, but is also able to sort and collect drops of interest, containing viable and healthy cells, which can be used for further experimentation. In this study, we used isogenic algal cells that were first synchronized in mixotrophic growth conditions. We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers) and a significant subpopulation of slowly dividing cells (slow-growers). These observations suggest that algal cells from an isogenic population may be present in either of two states, a state of restricted division and a state of active division. When isogenic cells were allowed to propagate for about 1000 generations on solid agar plates, they displayed an increased heterogeneity in their growth dynamics. Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times. Moreover, populations dividing with the same growth-rate often reached different cell numbers in stationary phase, suggesting that the progenitor cells differed in the number of cell divisions they could undertake. We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

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Growth curves of Chlamydomonas in droplets containing a low initial cell number (average of 0.16–0.19 cells/drop).Growth curves for Samples A from (a) WT222+, (b) WT11+ and (c) WTS24- or for Samples B from (d) WT222+, (e) WT11+ and (f) WTS24-. Selection of a set of growth curves showing the diversity in growth-rates and final algal yields, observed in Samples B batches for (g) WT222+ (h) WT11+ and (i) WTS24-. Note the sole existence of the two categories of slow- and fast-growers for Sample A batches, and the many intermediate growth phenotypes observed for Sample B batches.
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pone.0118987.g005: Growth curves of Chlamydomonas in droplets containing a low initial cell number (average of 0.16–0.19 cells/drop).Growth curves for Samples A from (a) WT222+, (b) WT11+ and (c) WTS24- or for Samples B from (d) WT222+, (e) WT11+ and (f) WTS24-. Selection of a set of growth curves showing the diversity in growth-rates and final algal yields, observed in Samples B batches for (g) WT222+ (h) WT11+ and (i) WTS24-. Note the sole existence of the two categories of slow- and fast-growers for Sample A batches, and the many intermediate growth phenotypes observed for Sample B batches.

Mentions: Fig. 5a shows the millifluidic growth curves of sample A of the WT222+ strain, for 0.29 inoculated cells on average per drop, meaning that the train of drops is comprised mainly of a mix of empty drops and drops inoculated with only one cell. It is apparent from the growth curves over 140 hours for Sample A in Fig. 5a, that there are two main distinct algal populations: in 60% of the occupied drops the final algal yield was of 800–1200 cells/droplet, whereas in 30% of the occupied drops, the final yield was below 100 cells/droplet, i.e. about 6.105 cells.mL-1. This latter concentration is typical of Chlamydomonas bulk cultures in early exponential phase in common laboratory conditions, and is far from the 1–2.107 cells.mL-1 usually reached in the stationary phase [59,79]. Fig. 5b and 5c confirm this main observation, using Sample A from WT11+ and from WTS24-. The corresponding millifluidic growth curves respectively correspond to 0.16 and 0.19 inoculated cells per drop on average. In both cases, two main distinct algal populations were again observed: a majority of drops yield high final cell concentrations (1200–2000), while a minor subpopulation yields a much lower final algal concentration, below 200 cells.


A millifluidic study of cell-to-cell heterogeneity in growth-rate and cell-division capability in populations of isogenic cells of Chlamydomonas reinhardtii.

Damodaran SP, Eberhard S, Boitard L, Rodriguez JG, Wang Y, Bremond N, Baudry J, Bibette J, Wollman FA - PLoS ONE (2015)

Growth curves of Chlamydomonas in droplets containing a low initial cell number (average of 0.16–0.19 cells/drop).Growth curves for Samples A from (a) WT222+, (b) WT11+ and (c) WTS24- or for Samples B from (d) WT222+, (e) WT11+ and (f) WTS24-. Selection of a set of growth curves showing the diversity in growth-rates and final algal yields, observed in Samples B batches for (g) WT222+ (h) WT11+ and (i) WTS24-. Note the sole existence of the two categories of slow- and fast-growers for Sample A batches, and the many intermediate growth phenotypes observed for Sample B batches.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4356620&req=5

pone.0118987.g005: Growth curves of Chlamydomonas in droplets containing a low initial cell number (average of 0.16–0.19 cells/drop).Growth curves for Samples A from (a) WT222+, (b) WT11+ and (c) WTS24- or for Samples B from (d) WT222+, (e) WT11+ and (f) WTS24-. Selection of a set of growth curves showing the diversity in growth-rates and final algal yields, observed in Samples B batches for (g) WT222+ (h) WT11+ and (i) WTS24-. Note the sole existence of the two categories of slow- and fast-growers for Sample A batches, and the many intermediate growth phenotypes observed for Sample B batches.
Mentions: Fig. 5a shows the millifluidic growth curves of sample A of the WT222+ strain, for 0.29 inoculated cells on average per drop, meaning that the train of drops is comprised mainly of a mix of empty drops and drops inoculated with only one cell. It is apparent from the growth curves over 140 hours for Sample A in Fig. 5a, that there are two main distinct algal populations: in 60% of the occupied drops the final algal yield was of 800–1200 cells/droplet, whereas in 30% of the occupied drops, the final yield was below 100 cells/droplet, i.e. about 6.105 cells.mL-1. This latter concentration is typical of Chlamydomonas bulk cultures in early exponential phase in common laboratory conditions, and is far from the 1–2.107 cells.mL-1 usually reached in the stationary phase [59,79]. Fig. 5b and 5c confirm this main observation, using Sample A from WT11+ and from WTS24-. The corresponding millifluidic growth curves respectively correspond to 0.16 and 0.19 inoculated cells per drop on average. In both cases, two main distinct algal populations were again observed: a majority of drops yield high final cell concentrations (1200–2000), while a minor subpopulation yields a much lower final algal concentration, below 200 cells.

Bottom Line: We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers) and a significant subpopulation of slowly dividing cells (slow-growers).Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times.We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Colloïdes et Matériaux Divisés, Institute of Chemistry, Biology and Innovation ESPCI ParisTech/CNRS UMR 8231/PSL* Research University, Paris, France.

ABSTRACT
To address possible cell-to-cell heterogeneity in growth dynamics of isogenic cell populations of Chlamydomonas reinhardtii, we developed a millifluidic drop-based device that not only allows the analysis of populations grown from single cells over periods of a week, but is also able to sort and collect drops of interest, containing viable and healthy cells, which can be used for further experimentation. In this study, we used isogenic algal cells that were first synchronized in mixotrophic growth conditions. We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers) and a significant subpopulation of slowly dividing cells (slow-growers). These observations suggest that algal cells from an isogenic population may be present in either of two states, a state of restricted division and a state of active division. When isogenic cells were allowed to propagate for about 1000 generations on solid agar plates, they displayed an increased heterogeneity in their growth dynamics. Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times. Moreover, populations dividing with the same growth-rate often reached different cell numbers in stationary phase, suggesting that the progenitor cells differed in the number of cell divisions they could undertake. We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

Show MeSH